1. Field of the Invention
The invention relates generally to the field of histology. In particular, the present invention is directed to an apparatus and method for automatically producing tissue slides.
2. Description of Related Art
Histology, the study of the groups of tissues found in most multicellular plants and animals, often requires that the plant or animal tissue be provided in thin sections for study. One known technique to create thin sections of tissue is through the use of a microtome. The function of the microtome is simplistic and very much like that of any mechanical slicer which carves a pre-set measured slice and retains and advances that unit of measure for each subsequent cutting cycle. The cutting results are very much dependent upon a technician's practical skill.
Typically, tissue samples for general pathology examination are surrounded by a support medium and manually molded into standardized blocks. Paraffin is a known and commonly-used as a support medium. TissueTek O.C.T., manufactured by Sakura Finetek and available from VWR Scientific, West Chester, Pa., may also be used as a support medium. The optimum cutting temperature (O.C.T.) formulation of water-soluble glycols and resins in TissueTek O.C.T. provides a convenient specimen matrix for cryostat sectioning at temperatures of −10° C. and below. Resins and polymers, such as methacrylates, may also be used as support media. Although resins and polymers are primarily used for electron microscopy, they can also be used for standard microscopy with some adjustments for hardness of the final product. Suitable resins and polymers, including Araldite 502 Kit, Eponate 12™ Kit, and Glycol Methacrylate (GMA) Kit, are available from Ted Pella, Inc., Redding, Calif.
The tissue sample is manipulated in a metal support medium block mold by the technician so as to locate it close to what will become the top center surface of the support medium block. This top surface then becomes the working surface of the sample block. The sample block preparation process will be described using paraffin as the support medium.
As the tissue sample is being located and set within the support medium block mold, melted paraffin is added. The paraffin cools and hardens inside the block mold; and as it does, a plastic holder is inserted. In the hardening process, the bottom half of the paraffin block becomes locked within the frame of the plastic holder which becomes the base of the sample block. A technician then transfers the sample block to a microtome to prepare very thin sections of the tissue sample for later observation.
The prepared sample blocks are manually locked into an adjustable clamp assembly of the microtome. The technician can make limited manual adjustments to the clamp assembly in order to present the sample block's working surface to the microtome's cutting blade at an angle judged to be most favorable to yielding the best possible sample slice. The clamp adjustment for a given sample block is a subjective choice based on the judgement and experience of the technician.
The technician can make additional microtome adjustments that pre-set the starting lineal distance between the clamp assembly and the assembly holding the cutting blade, as well as the thickness of the sample slice each time the clamp assembly is cycled by moving it past the blade assembly. There is also limited adjustment of the angle of the cutting blade as presented to the clamp assembly; however, the optimum setting of the blade is typically about 45 degrees from vertical.
One sample slice is the product of one microtome cycle. Once a technician selects the orientation of the sample block's working surface to the cutting blade assembly's cutting blade, several cutting cycles are required to advance the cutting blade into the block to a working surface depth, where the slices then taken are suitable for slide application. These slices may be referred to as histologic grade slices, which are slices that a pathologist or other professional would find of acceptable quality for examination purposes. In particular, histologic grade slices are slices of a relatively uniform thickness without any significant tears, folds, wrinkles, or contamination from unrelated specimens or other artifacts. Histologic grade slices are substantially flat and smooth.
When a suitable sample depth is acquired, the technician may cycle the clamp (with the sample block locked within) one to several times to obtain a series of histologic grade slices. After each slice, a mechanism in the microtome advances the block toward the knife by the desired slice thickness, which may be set by the technician. A typical slice thickness is about 4 to 8 microns, but may vary from 1 to more than 10 microns.
The technician places the slices onto the surface of warm water, located in a pan next to the microtome. The tissue sample, still secured in its paraffin matrix, floats on the surface of the water. The warmth of the water softens the paraffin, thereby removing wrinkles or other distortions in the slice. The technician may also move a sample slice about on the surface of the water to facilitate this result.
Once the technician judges that the floating tissue/paraffin matrix sample has the desired flat form, the technician brings a receiving medium, such as a microscope slide, up under the portion of the floating sample slice that captures the full tissue specimen. Repetitive success in this, again, depends upon the skill of the technician. The slide with sample is then set aside for drying, staining and evaluation, or for various molecular or histochemical analysis.
Occasionally, such as during performance of surgical procedures, it may be necessary to get a rapid diagnosis of a pathologic process. For example, a surgeon may want to know if the margins of his resection for a malignant neoplasm are clear before closing, or an unexpected disease process may be found and require diagnosis to decide what to do next, or it may be necessary to determine if the appropriate tissue has been obtained for further workup of a disease process. This may accomplished through use of a frozen section.
Frozen sections are performed with an instrument called a cryostat. The cryostat is essentially a refrigerated box containing a microtome. The temperature inside the cryostat is about −20 to −30° C. The piece of tissue to be studied is snap frozen in a cold liquid or cold environment (−20 to −70° C.). TissueTek O.C.T. would be a suitable support medium for frozen sections. The freezing makes the tissue solid enough to section with a microtome. The tissue sections are cut and picked up on a glass slide. The sections are then ready for staining.
Although this process is effective, it is very time consuming and very dependent on the skill of the technician. As hospitals and laboratories seek to lower operating costs, they will seek to find more efficient ways to accomplish inefficient tasks. Also, it is desirable to reduce the variation in sample preparation that may occur between different technicians.